Hematopoietic stem cells (HSCs) are defined by their unique ability to differentiate into all mature blood cell lineages while at the same time regenerating themselves, in a process termed self-renewal, to sustain hematopoiesis throughout mammalian life. A model explaining the nature of HSC divisions proposes that the mode of a division of a stem cell (symmetric versus asymmetric) may determine the cell fate of the resulting two daughter cells, with a symmetric division resulting in daughter cells with simila regenerative potential and an asymmetric division leading to daughter cells with a dissimilar potential. Terminologies like polarity and division symmetry have already been previously discussed in other stem cell systems, including hematopoiesis. The molecular mechanisms of the triad HSC polarity - division symmetry - cell fate though has eluded the HSC field to date and which factors instruct this process is not known. The markers of polarity associated with division asymmetry are also ill-defined. We propose to investigate a molecular link, i.e. Cdc42 activity, in the stem cell polarity - mode of division - cell fate relationship. A new concept that Cdc42 activity and cell polarity represent a dial-able threshold in a bell shaped manner in determining stem cell fate (i.e. high and low Cdc42 activity could both cause a loss of polarity and affect the mode of the cell division and fate), is proposed. A rigorous examination of the influence of Cdc42 mediated polarity/apolarity on the mode of stem cell division and daughter cell fate will strongly benefit the stem cell field and will impact the therapy of blood diseases such as bone marrow failure and aging. We propose three specific aims: First, we will examine the potentially causal relationship between cell polarity and the mode of HSC division by a series of in vitro and in vivo cell polarization and division symmetry analysis at single cell resolution. Second, we will define the role of Cdc42 in regulating the ratio between polar and apolar HSCs and consequently the cell fate. Third, we will determine the signaling pathway from HSC polarity into divisional asymmetry by examining non-canonical Wnt5a regulated Cdc42 activity and subsequent actomyosin and JNK/p38 signaling in polarity and division symmetry of HSCs. The studies will use state of the art immunofluorescence, chemical biology, mouse genetics, and stem cell methodologies like single cell transplants to define the molecular basis of cell polarity and divisional symmetry and their causal role in determining HSC cell fate.